Lipidemic, glycemic and organ protective actions of tea seed oil in rats fed with high fat
and high carbohydrate diet
Warinna Pinthong1,*,#, Thamolwan Suanarunsawat2, Watcharaporn Devakul Na Ayutthaya3
1
Biomedical Sciences Graduate Program, Department of Medical Sciences, Faculty of
Science, Rangsit University, Thailand
2
Physiology Unit, Department of Medical Sciences, Faculty of Science, Rangsit University,
Thailand
3
Pharmacology and Toxicology Unit, Department of Medical Sciences, Faculty of Science,
Rangsit University, Thailand
*,#e-mail: warinna_pinthong@hotmail.com
Abstract
The present study was conducted to investigate the lipidemic, glycemic and organ
protective effects of tea seed oil (TSO) in rats fed with high fat and high carbohydrate
(HFHC) diet. Three groups of male Wistar rats were used including normal control group,
group fed with HFHC diet (cholesterol + lard + fructose) for three months, and HFHC group
treated with TSO. At the end of the experiment, serum lipid profile, blood glucose, oral
glucose tolerance test (OGTT), serum AST, ALT, LDH, CK-MB, creatinine and BUN were
determined. Individual fatty acid contents in TSO were assayed by Gas chromatography. The
results showed that oleic acid was the primary fatty acid in TSO (83.36%). HFHC diet
increased serum lipid profile and atherogenic index (AI). The high serum levels of lipid
profile and AI were decreased in HFHC rats treated with TSO. The elevation of area under
the curve of OGTT was alleviated by TSO. TSO also normalized the high serum levels of
AST, ALT, LDH, CK-MB, creatinine and BUN. It can be concluded that TSO was able to
decrease the high serum levels of lipid profile and high blood glucose level of OGTT in rats
fed with HFHC, indicating its therapeutic potency to prevent atherosclerosis and
hyperglycemia. It also protects liver, heart and kidney in HFHC. Oleic acid content in TSO
might be responsible for these activities.
Keywords: tea seed oil, high fat and high carbohydrate diet, liver function, cardiac function,
fructose
Introduction
Hyperlipidemia and hyperglycemia are widely known to be the major risk factor for
the development of atherosclerosis, ultimately causes coronary artery disease [1, 2]. The
modern life style with high fat and high carbohydrate diet and less physical activity are the
significant causes of hyperlipidemia and hyperglycemia. Hyperlipidemia and hyperglycemia
have been found to induce oxidative stress in various organs such as liver, heart and kidney
[3, 4]. Since synthetic hyperlipidemia and hyperglycemia drugs cause several adverse effects
especially liver damage, searching for natural products that have less or no side effects has
been increasing. Among many kinds of medicinal plants available in Thailand, Camellia
oleifera Abel. oil (tea seed oil, TSO) is promising for hyperlipidemia and hyperglycemia
drugs since it has several outstanding properties such as high smoking point, stable, and high
nutritional contents, especially Vitamin E, and high unsaturated fat which is useful for human
foods [5]. TSO is extensively used as cooking oil in several countries including China and
Thailand. Tea seed oil has been shown to lower serum lipid profile [6, 7] and protected vital
organs in normal and several stress conditions as reported earlier [8, 9, 10]. However, no
experimental study to elucidate its effects on serum lipid profile and blood glucose in subjects
fed with high fat and high carbohydrate diet has been reported. Therefore, the present study
was conducted to investigate its lipidemic, glycemic and organ protective effects in rats fed
with high fat and high carbohydrate diet.
Methodology
1. Animal preparation
Male Wistar rats weighing between 180-220 g from the National Laboratory Animal
Centre, Mahidol University, Salaya, Nakornprathom, Thailand were used in the study. The
animals were housed in the animal facility of the Faculty of Science, Rangsit University
under standard conditions of temperature (25±2°C), 50-60% of humidity and 12 h/12 h
light/dark cycles. Food and water were given ad libitum. High fat and high carbohydrate
(HFHC) diet was prepared by adding 2 g% cholesterol powder along with 10% lard in normal
diet, and 60% fructose powder mixed in drinking water. Body weight and food consumption
were recorded weekly in all groups. Three groups of seven rats each were fed on the
following diet for three month: group I, normal diet for a control; group II, a high fat and high
carbohydrate (HFHC) diet; and group III, HFHC diet treated with TSO was orally
administered once a day at the dose of 5.69 g/kg BW/day for the last three months. This dose
has been chosen because of its anti-oxidative effect to protect liver against CCl4 [10]. In
addition, it is the average dose that showed blood lipid lowering in previous reports [7, 11].
The tea seed oil was bought from the local market.
2. Experimental protocol
2.1 Effect of tea seed oil on blood glucose and oral glucose tolerance tests in rats fed with
HFHC diet
After 3 months of normal or HFHC diet feeding, the rats were fasted overnight. Blood
was collected from rats’ tail to determine fasting blood glucose (FBG) using automatic
glucometer (One Touch Ultra, Lifescan, CA, USA). Oral glucose tolerance test was evaluated
by feeding 2 g/kg BW of 50% glucose solution. After glucose loading, blood was collected
from the tail to determine blood glucose every 30 min for 120 min and the rats were
anesthetized by intraperitoneal injection with zolitil (40 mg/kg BW) plus xylazine (3 mg/kg
BW).
2.2 Effect of tea seed oil on serum lipid profile in rats fed with HFHC diet
At the end of the experiment, the rats were fasted overnight, and were anesthetized by
intraperitoneal injection with sodium pentobarbiturate (60 mg/kg BW). Blood was collected
from abdominal vein. Serum was separated by refrigerated centrifuge at 3000 rpm, 4°C for 5
min for determination of serum lipid profile including total cholesterol (TC), triglyceride
(TG), high density lipoprotein cholesterol (HDL-C) and low density lipoprotein cholesterol
(LDL-C).
2.3 Organ protective effect of tea seed oil in rats fed with HFHC diet
At the end of the experiment, the rats were fasted overnight, and were anesthetized by
intraperitoneal injection with sodium pentobarbiturate (60 mg/kg BW). Blood was collected
from abdominal vein. Serum was separated by refrigerated centrifuge at 3000 rpm, 4°C for 5
min. Liver damage was evaluated by determination of serum alanine aminotransferase
(ALT), aspartate aminotransferase (AST). Cardiac damage was also assessed by
determination of serum lactate dehydrogenase (LDH) and creatine kinase MB subunit (CKMB). Renal function was evaluated by determination of serum creatinine and blood urea
nitrogen (BUN).
2.4 Biochemical Assay
The total serum levels of total cholesterol, triglyceride and HDL-C were assayed by
using an enzymatic kit (Gesellschaft für Biochemica und Diagnostica GmbH, Germany).
LDL-C was calculated by using the equation: LDL-C = [TC-(HDL-C)]-(triglyceride/5). The
serum levels of AST, ALT, LDH, and CK–MB were measured by using an enzymatic kit
(Randox Laboratories, UK). Total cholesterol and triglyceride contents in the liver and feces
were determined using enzymatic kit.
3. Statistical Analysis
All values were presented as means ± SEM. The results were analyzed by ANOVA.
Duncan multiple rank test was performed to determine statistical significance among groups
by using SPSS software version 11.5. Significant difference was accepted at p < 0.05.
Results
Table 1 and Figure 1 showed that basal blood glucose was slightly increased in rats
fed with HFHC diet but the level was not significantly different from normal rats. TSO
treatment slightly decreased basal blood glucose but the level was not significantly different
from HFHC rats.
Table 1. Basal blood glucose in normal rats, HFHC rat and HFHC rats treated with TSO
Group
Blood glucose (mg/dl)
84.43 ± 3.14a
control
94.71 ± 2.85a
HFHC
90.71 ± 4.13a
HFHC+TSO
Values are expressed as mean ± SEM of seven rats per group. Values with different
superscripts in the same row are significantly different at p < 0.05. HFHC: high fat and high
carbohydrate.
Figure 1. Basal blood glucose in normal rats, HFHC rat and HFHC rats treated with TSO
Values are expressed as mean ± SEM of seven rats per group. Values with different
superscripts in the same row are significantly different at p < 0.05. HFHC: high fat and
high carbohydrate.
In contrast, area under the curve (AUC) of oral glucose tolerance test (OGTT) was
significantly increased in rats fed with HFHC diet as compared to normal rats. The high level
of AUC was significantly lowered in HFHC rats treated with TSO (Table 2 and Figure 2).
Table 2. Area under the curve of oral glucose tolerance test (OGTT) in normal rats,
HFHC rat and HFHC rats treated with TSO
Group
Area under curve of blood glucose
(mg/120 min)
544.43 ± 12.76a
control
636.79 ± 16.45b
HFHC
591.07 ± 8.56c
HFHC+TSO
Values are expressed as mean ± SEM of seven rats per group. Values with different
superscripts in the same row are significantly different at p < 0.05. HFHC: high fat and
high carbohydrate.
Figure 2. Oral glucose tolerance test in normal rats, HFHC rat and HFHC rats treated with TSO
The serum levels of total cholesterol, triglyceride, HDL-C, LDL-C and atherogenic
index (AI) were shown in the Table 3 and Figure 3. Three months of HFHC diet feeding
significantly increased serum total cholesterol, triglyceride, LDL-C and AI whereas HDL-C
was slightly decreased. TSO treatment significantly decreased the levels of total cholesterol,
triglyceride, LDL-C, and atherogenic index (AI) in HFHC rats.
Table 3. Serum total cholesterol, triglyceride, HDL-C, LDL-C, atherogenic index (AI) in normal rats, HFHC
rat and HFHC rats treated with TSO
Group
Total cholesterol
(mg/dl)
Triglyceride
(mg/dl)
HDL-C
(mg/dl)
LDL-C
(mg/dl)
Atherogenic
Index
control
HFHC
HFHC+TSO
54 ± 5.67a
157 ± 8.64b
120 ± 9.16c
29 ± 3.57a
50 ± 5.27b
21 ± 2.12a
21.19 ±1.98 a
17.49 ± 1.57a
18.92±1.74a
26.97±3.49a
130.04±8.92b
96.86±10.11c
1.54±0.11a
8.49±1.01b
5.86±1.07c
Values are expressed as mean ± SEM of seven rats per group. Values with different superscripts in each
group are significantly different at p < 0.05.
Figure 3. Serum total cholesterol, triglyceride, HDL-C, LDL-C (Left), and atherogenic index (Right) in
normal rats, HFHC rat and HFHC rats treated with TSO
The serum levels of AST, ALT, LDH, CK-MB, creatinine, and BUN were shown in
Table 4. Three months of HFHC diet feeding significantly increased serum levels of AST,
ALT, LDH, CK-MB, creatinine, and BUN. The high levels of AST, ALT, LDH, CK-MB,
and BUN were significantly decreased in HFHC rats treated with TSO.
Table 4. Serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), creatine
kinase MB subunit (CK-MB), lactate dehydrogenase (LDH), creatinine and BUN in normal rat, HFHC rat
and HFHC rat treated of TSO
Group
Serum AST
(U/L)
Serum ALT
(U/L)
CK-MB
(U/L)
LDH
(U/L)
Creatinine
(mg/dl)
BUN
(mg/dl)
Control
HFHC
HFHC+TSO
83 ± 9a
181 ± 12b
119 ± 11c
33 ± 3a
142 ± 14b
54 ± 6a
83 ± 9a
181 ± 12b
119 ± 11c
33 ± 3a
142 ± 14b
54 ± 6a
581 ± 40a,b
757 ± 58b
536 ± 79a
454 ± 43a
579 ± 23b
415 ± 43a
Values are expressed as mean ± SEM of seven rats per group. Values with different superscripts in each
column are significantly different at p < 0.05.
Discussion and Conclusion
It has been widely known that elevation of hyperlipidemia and hyperglycemia can
lead to atherosclerosis; blood supply to the organs gradually diminishes until organ function
becomes impaired. Several lines of evidence show that the improvement and incidence of
atherosclerosis and coronary artery disease are associated with lowering hyperlipidemia [1, 2]
and hyperglycemia [3]. To treat hyperlipidemia and hyperglycemia, extensive interventions
are recommended, including diet control and exercise. Hyperlipidemia and hyperglycemia
drugs can cause several adverse effects especially liver damage indicating the need to find
other safer yet effective alternatives [12, 13]. Therefore, herbal medicines might be an
effective, safe, and low-cost therapy. Camellia oleifera Abel. oil (TSO) is very promising
due to its therapeutic potency [5]. The present study shows that three months of HFHC diet
feeding raised the basal blood glucose but the level was not significantly different from
normal rats. TSO treatment slightly decreased basal blood glucose but the level was not
significantly different from HFHC rats. In contrast, AUC of OGTT was significantly
increased in rats fed with HFHC diet. The substantial increase in the amount of dietary
fructose consumption from high intake of sucrose has been linked with a rise in obesity and
metabolic disorders. A high flux of fructose to the liver perturbs glucose metabolism and
glucose uptake pathways, leads to a significantly enhanced rate of de novo lipogenesis and
triglyceride synthesis. Because of its lipogenic properties, excess fructose in the diet can
cause glucose and fructose malabsorption, and greater elevations in TG and cholesterol.
These metabolic disturbances appear to underlie the induction of insulin resistance or
impairment of insulin action commonly observed with high fructose feeding in both humans
and animal models and then finally results hyperglycemia [7, 10, 11]. Providing fructose
with honey, which is naturally rich in antioxidant substances, prevented both the oxidative
stress induced by fructose and the reduction of insulin sensitivity [12]. TSO significantly
decreased the high level of AUC in HFHC diet, this implies that TSO was able to stimulate
insulin secretion or improves insulin action which then eventually prevents the occurrence of
anti-hyperglycemic [13]. Research studies that provide evidence TSO effect of decreased
total cholesterol, triglyceride, phospholipid and lipid profile in the liver and heart in human
and rat [14, 15]. In the present study, HFHC diet also increased serum lipid profile and
atherogenic index (AI). Hyperlipidemia is an important factor in development of
atherosclerosis and finally coronary artery disease. HFHC diet caused oxidative stress as
shown by marked increment in the levels of thiobarbituric acid reactive substance (TBARS)
and reduction of reduced glutathione content, decreased activities of SOD, CAT and GPx in
the liver, heart and kidney [16, 17]. TSO treatment attenuated the high serum lipid profile
and atherogenic index (AI). This is supported by the increased activities of SOD, CAT and
GPx in the liver, heart and kidney. These results suggest that TSO could be effective to
alleviate atherosclerosis which eventually prevents the occurrence of coronary artery disease
[18]. Free radical-induced lipid peroxidation or oxidative stress has been shown to
participate in the pathogenesis of several diseases [19]. In addition, it was shown that
activities of antioxidant enzymes increased in both the liver, kidney and cardiac tissues of rats
after TSO administration [20, 21]. The liver, heart and kidney are widely known to be the
major risk organs for hyperlipidemia and hyperglycemia. In the present study, liver, heart and
kidney were damaged as shown by the increased of serum levels of AST, ALT, LDH, CKMB, creatinine, and BUN in HFHC rats (Table 4). HFHC diet feeding induces lipid
peroxidation or oxidative stress to injure the liver, heart and kidney. TSO decreased all these
serum marker enzymes. In previous research, TSO showed higher antioxidant enzyme (SOD)
activity served as anti-active oxygen species in body tissues. On the other hand, TSO can
increase antioxidant enzyme activity [22]. Therefore, TSO may protect the liver, heart and
kidney from HFHC diet by which antilipidemic, antiglycemic and antioxidative actions. In
previous studies, qualitative analysis of the fatty acid composition of TSO shows that the
main fatty acid compositions are relatively stable. It is mainly composed of oleic acid has the
effect of lipid-lowering in the blood, reduce lipid peroxidation and increase capacity of
antioxidants in the plasma and liver in rat [23, 24]. It has been shown that oleic acid
decreased hepatic lipid content whereas increased fecal bile acids excretion without
significant change of fecal lipid excretion [25]. In this study, oleic acid (83.36%) was the
primary fatty acid in TSO. These results indicate that oleic acid containing in TSO may play
important role for these effects.
In summary, consumption of the HFHC diet feeding raised blood glucose, serum lipid
profile and AI. The liver, heart and kidney functions were also damaged. TSO was able to
decrease the high serum levels of lipid profile and blood glucose in rats fed with HFHC,
indicating its therapeutic potency to prevent atherosclerosis and hyperglycemia. TSO also
protects liver, heart and kidney. Oleic acid containing in TSO might be responsible for
hypolipidemic, hypoglycemic and organ protective activities.
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Acknowledgement: This work was supported by Rangsit University, Patumthani, Thailand